Affinity labeling techniques were employed to identify anc characterize the plasma membrane receptors for 3,3 feet, 5-triiodo-L-thyronine (T3) in cultured Swiss 3T3-4 mouse fibroblasts, GH3 rat pituitary tumor cells and human epithelioid carcinoma A431 cells. A major specifically labeled protien with an apparent molecular mass of 55 kDalton (kDal) was detected in three cell lines. One-dimensional peptide mapping showed there are structural similarities in the 55-kDal protein from three different species. Thus, the plasma membrane T3 receptors are highly conserved. Using 125I-labeled L-thyroxine ([125I]T4), the binding and uptake of T4 in cultured GH3 cells and Swiss 3T3-4 cells were shown to be saturable and specific. These results showed that the uptake of T4 is receptor-mediated. Using affinity labeling and peptide mapping techniques, plasma membrane T3 and T4 receptors were shown to have structural similarities. The results form equilibrium binding studies indicate that one plasma membrane thyroid hormone receptor mediates the uptake of both T3 and T4 into cells. Using electron spin resonance (ESR), the dynamic interactions of thyroid hormones with liposomes derived from L-Alpha-dimyristoyl-phosphatidylcholine (DMPC) and plasma membranes of GH3 cells was studied. Using the motional narrowing formalisms, the rotational correlation times of spin-labeled T4 in DMPC vesicles at 31 degrees C were estimated at 3.1 x 10 to the -3 and 4.8 x 10 to the -9 sec for the linear term and the quadratic term, respectively. The rate of lateral diffusion of spin-labeled T4 in DMPC was approximately 5.2 x 10 to the -8 cm2/sec as determined by the ESR line-broadening method. These results indicate that spin-labeled thyroxine diffuses freely in the DMPC matrix. Studies using spin-labeled T3 (SL-T3) in DMPC vesicles gave similar results. However, analyses of the ESR spectrum from the binding of SL-T3 to the plasma membranes of GH3 cells indicate that Sl-T3 is highly immobilized. These results suggest that SL-T3 binds to a protein component. The interaction of SL-T3 with plasma membranes of GH3 cells is being characterized.